Even though C. burnetii has been recognized as the Q fever agent since the 1930s, we still have an incomplete understanding of host cell entry, disease pathogenesis or how the agent avoids immune clearance. Toll-like receptor (TLR) pathways have been hypothesized to play a role in bacterial uptake and clearance, but our understanding of TLRs in Q-fever is incomplete. Though a few studies of C. burnetii infection of the peritoneum in TLR deficient animals have been published, to date there have been no reports using TLR deficient animals and the natural route of infection via the lung. Lung TLR usage is unique, thus these studies are required to gain a truly relevant picture of the role of TLRs in C. burnetii infection. In preliminary studies we have detected differences in bacterial clearance in TLR2, TLR4 and MyD88 (Key inflammatory signaling molecule for TLRs 2&4) deficient animals following pulmonary challenge, some of which (differences) are contrary to what others have defined following peritoneal infection. We propose to expand on and confirm these preliminary observations. Another deficiency in our understanding of the role of TLRs in C. burnetii infection is whether TLR function is most relevant to bone-marrow derived macrophages, or whether TLRs on non-macrophages or even non-bone-marrow derived cells significantly contribute to host responses against the bacterium. Of particular interest is the potential role of TLR recogntion/function by mast cells in C. burnetii pathogenesis. These issues will also be examined in the proposed studies. Finally, an innovative component of the proposed studies will involve use of novel, human TLR4/MD2 transgenic mice to compare human and mouse TLR4 responses to C. burnetii in vivo. The specific hypothesis to be tested in this proposal is: TLR signaling is important in the host/C. burnetii interaction in the lung. This hypothesis will be addressed in the following Specific Aims. Aim 1: Compare C. burnetii infection in wild type, and TLR- and MyD88 deficient mice following lung challenge. Aim 2: Determine the importance of TLRs on the responses of hematopoietic and non-hematopoietic cells, including the role of mast cells following C. burnetii infection in the lung. Aim 3: Compare C. burnetii infection response in human TLR4/MD2 transgenic and mouse TLR4 competent mice.